Pseudo-viral particles and uses of same

Abstract

The present invention relates to a type I or II transmembrane fusion protein comprising, successively: a) optionally, a signal peptide; b) a protein or a peptide of interest; c) a coiled-coil domain; and d) a domain for anchoring in the plasma membrane, consisting of a transmembrane segment and a cytosolic segment. It also relates to the virus-like particles obtained with this fusion protein.

1 . A virus-like particle comprising:
 an envelope consisting of a plasma membrane of which at least one portion is typical of lipid rafts; and   at least one type I or II transmembrane fusion protein anchored in said membrane, said fusion protein comprising the following fragments, successively:   b) a protein or a peptide of interest;   c) a coiled-coil domain or oligomerization sequence, which does not originate from a virus; and   d) a domain for anchoring in the plasma membrane, consisting of a transmembrane segment and a cytosolic segment, preferably a domain for anchoring in a plasma membrane of which at least one portion is typical of lipid rafts, fragments b) and c) being exposed at the surface of the virus-like particle.   
     
     
         2 . The virus-like particle according to  claim 1 , wherein a linker is present between fragments b) and c), and/or between fragments c) and d). 
     
     
         3 . The virus-like particle according to  claim 1 , wherein:
 b) the protein or the peptide of interest is chosen from:
 allergens and fragments thereof, 
 cell surface proteins and fragments thereof, 
 proteins and peptides that are accumulated in chronic or neurodegenerative diseases, 
 proteins and peptides involved in hypertension, 
 immunoglobulins and fragments thereof, 
 cytokines and fragments thereof, and 
 hormones and fragments thereof; 
   c) the coiled-coil domain is chosen from SEQ ID NO: 24, SEQ ID NO: 27, SEQ ID NO: 28, SEQ ID NO: 29, SEQ ID NO: 33 and SEQ ID NO: 30; and   d) the anchoring domain is chosen from the anchoring sequence of the H5N1 influenza virus H5 hemagglutinin (SEQ ID NO: 26) and the anchoring sequence of the PDLP1 protein (SEQ ID NO: 31).   
     
     
         4 . The virus-like particle according to  claim 1 , wherein the protein or the peptide of interest is chosen from allergens and fragments thereof. 
     
     
         5 . A type I or type II transmembrane fusion protein comprising the following fragments, successively:
 a) optionally, a signal peptide;   b) a protein or a peptide of interest;   c) a coiled-coil domain or oligomerization sequence, which does not originate from a virus; and   d) a domain for anchoring in the plasma membrane, consisting of a transmembrane segment and a cytosolic segment, preferably a domain for anchoring in a plasma membrane of which at least one portion is typical of lipid rafts.   
     
     
         6 . The transmembrane fusion protein according to  claim 5 , wherein the protein or the peptide of interest is chosen from allergens and fragments thereof. 
     
     
         7 . A nucleic acid encoding a fusion protein as claimed in  claim 5 . 
     
     
         8 . A host cell or a vector comprising at least one nucleic acid according to  claim 7 . 
     
     
         9 . A method of treatment comprising a step of administering, to a patient in need thereof, a therapeutically effective amount of the virus-like particle  claim 1 . 
     
     
         10 . A method of allergen immunotherapy comprising a step of administering, to a patient in need thereof, a therapeutically effective amount of the virus-like particle according to  claim 1 . 
     
     
         11 . A method for producing virus-like particles according to  claim 1 , comprising the expression of the nucleic acid as claimed in  claim 7  in eukaryotic cells. 
     
     
         12 . The method according to  claim 11 , characterized in that the eukaryotic cells are plant cells, and in that it comprises the following steps:
 a) transformation of agrobacteria with an expression vector comprising a nucleic acid as claimed in  claim 5  functionally linked to a strong promoter; and   b) transfection of the plant cells with the agrobacteria obtained in step a), said transfection comprising the following steps:   b1) culture of the plant cells, under aeroponic or hydroponic conditions, and under LED lighting, preferably for four to six weeks under hydroponic conditions,   b2) agroinfiltration of the plant cells obtained in b1) under vacuum, with the agrobacteria obtained in step a), preferably carried out under vacuum by Venturi effect,   b3) return to culture of the plant cells obtained in b2), typically for 3 to 6 days, in order to obtain the virus-like particles,   then extraction of the virus-like particles obtained and purification, in particular by enzymatic extraction.